Approximately 1% of the human genome appears to be of retroviral origin. The purpose of these "endogenous" retroviral elements in human development remains unknown. A small number of organs express endogenous retroviruses under physiologic conditions; the human placenta is one of these. The normal human placental villous cytotrophoblast expresses differentiation-related transcription of the env region of a single copy endogenous retroviral genome (ERV-3). The strong association of ERV-3 expression with normal trophoblast differentiation suggests a physiologic role for ERV-3 in placental development. In support of this hypothesis, are the following observations. 1) ERV-3 env mRNA and Env protein are increased in syncytiotrophoblast and in spontaneously differentiating mononuclear villous cytotrophoblast. 2) Differentiation-related increase in ERV-3 env mRNA is concurrent with increased b-hCG mRNA and protein and intercellular fusion. 3) Transfection of BeWo choriocarcinoma cells, a model for trophoblast differentiation, with the sense strand of ERV-3 env resulted in a stable transfectant in which most aspects of trophoblast differentiation occurred: decreased growth, morphologic differentiation, and increased b-hCG mRNA and hCG protein production. 4) Transfection with the antisense strand decreased b-hCG mRNA production when the cells were induced to differentiate. These preliminary data support a hypothesis that ERV-3 env expression is an important step in the pathway of normal trophoblast differentiation, potentially through regulation of hCG expression. The specific aim of this R03 application is to accumulate adequate preliminary data for a successful R01 application to test this hypothesis, The individual specific Aims are: Aim 1) is to develop reagents, e.g.,monoclonal antibody and cell lines transfected with inducible plasmid systems, for studying the role of ERV3 env in villous trophoblast differentiation; Aim 2) is to determine whether ERV-3 env expression is necessary for differentiation of freshly isolated villous cytotrophoblasts; and Aim 3) is to determine whether ERV-3 env expression is inducible by steroids. These specific aims were chosen to address issues raised in previously unsuccessful R01 applications.